While a host of molecular techniques are utilized by evolutionary developmental (evo-devo) biologists, tools for quantitative evaluation of morphology are still largely underappreciated, especially in studies on microscopic animals. Here, we provide a standardized protocol for geometric morphometric analyses of 2D landmark data sets using a combination of the geomorph and Morpho R packages. Furthermore, we integrate clustering approaches to identify group structures within such datasets. We demonstrate our protocol by performing exemplary analyses on stomatal shapes in the model nematodes Caenorhabditis and Pristionchus. Image acquisition for 80 worms takes 3–4 d, while the entire data analysis requires 10–30 min. In theory, this approach is adaptable to all microscopic model organisms to facilitate a thorough quantification of shape differences within and across species, adding to the methodological toolkit of evo-devo studies on morphological evolution and novelty.

尽管进化的（evo-devo）生物学家使用了许多分子技术，但用于形态学定量评估的工具仍大大不足，特别是在微观动物研究中。在这里，我们提供了用于结合使用geomorph和Morpho R软件包的2D地标数据集的几何形态分析的标准化协议。此外，我们集成了聚类方法以识别此类数据集中的组结构。我们通过对模型线虫Caenorhabditis和Pristionchus中的气孔形状进行示例性分析来证明我们的协议。80条蠕虫的图像采集需要3-4天，而整个数据分析需要10-30分钟。从理论上讲，这种方法适用于所有微观模型生物，以促进对物种内和物种间形状差异的彻底量化，从而增加了关于形态演变和新颖性的evo-devo研究的方法学工具包。